RNA interference refers to the process of sequence-specific post-transcriptional gene silencing in animals mediated by short interfering RNAs (siRNAs) (Fire et al. (1998) Nature 391:806-811). The corresponding process in plants is commonly referred to as post-transcriptional gene silencing (PTGS) or RNA silencing and is also referred to as quelling in fungi. The process of post-transcriptional gene silencing is thought to be an evolutionarily-conserved cellular defense mechanism used to prevent the expression of foreign genes and is commonly shared by diverse flora and phyla (Fire (1999) Trends Genet. 15:358-363). Such protection from foreign gene expression may have evolved in response to the production of double-stranded RNAs (dsRNAs) derived from viral infection or from the random integration of transposon elements into a host genome via a cellular response that specifically destroys homologous single-stranded RNA of viral genomic RNA. The presence of dsRNA in cells triggers the RNAi response through a mechanism that has yet to be fully characterized.
A new class of small RNA molecules is involved in regulating gene expression in a number of eukaryotic organisms ranging from animals to plants. These short RNAs or microRNAs (miRNAs; miRs) are 20-22 nucleotide-long molecules that specifically base-pair to target messenger-RNAs to repress their translation or to induce their degradation. Recent reports have identified numerous miRNAs from vertebrates, Caenorhabditis elegans, Drosophila and Arabidopsis thaliana (Bartel (2004) Cell 116:281-297; He and Hannon (2004) Nature Reviews Genetics 5:522-531).
Viruses such as Turnip Mosaic Virus (TuMV) and Turnip Yellow Mosaic Virus (TYMV) cause considerable crop loss world-wide and have serious economic impact on agriculture (Morch et al. (1998) Nucleic Acids Res 16:6157-6173; Skotnicki et al. (1992) Arch Virol 127:25-35; Tomlinson (1987) Ann Appl Biol 110:661-681). Most if not all plant viruses encode one or more proteins that are able to suppress the host's post-transcriptional gene silencing (PTGS) mechanism so as to ensure their successful replication in host cells. The PTGS is a mechanism that a plant host uses to defend against viruses by triggering breakdown of double stranded RNAs which are produced as intermediates in viral genome replication (Bernstein et al. (2001) Nature 409:363-366; Hamilton and Baulcombe (1999) Science 286:950-952; Zamore et al. (2000) Cell 31:25-33).
Reduction of the activity of specific genes (also known as gene silencing, or gene suppression), including virus genes, is desirable for several aspects of genetic engineering in plants. There is still a need for methods and constructs that induce gene suppression against a wide selection of target genes, and that result in effective silencing of the target gene at high efficiency.